Green fluorescent protein (GFP) derived from a jelly fish, Aequorea victoria, has many uses in biological systems. Recently, various GFP mutants that are changed in color, improved in folding characteristic, enhanced in luminance, and modified in pH sensitivity have been prepared based on random mutagenesis techniques and semi-rational mutagenesis techniques. Using genetic manipulation technology, another protein is fused to a fluorescent protein such as GFP, and monitoring of its expression and transport are being carried out.
As one of the most frequently used GFP mutants, yellow fluorescent protein (YFP) is listed. YFP shows fluorescence with the longest wavelength among jellyfish Aequorea GFP mutants. The values of ε and Φ of most YFPs are 60,000 to 100,000 M−3 cm−3 and 0.6 to 0.8, respectively (Tsien, R. Y. (1998). Ann. Rev. Biochem. 67, 509-544), and these values are comparable to those of common luminophores (fluorescein, rhodamine, etc.). Accordingly, improvement in absolute luminance of YFP is almost reaching the limit.
As other examples of the GFP mutants, there is cyan fluorescent protein (CFP), and enhanced cyan fluorescent protein (ECFP) is known. Further, red fluorescent protein (RFP) is also isolated from a Discoma sp., and DsRed is known. Thus, four typed of fluorescent proteins (green, yellow, cyan, and red) have been developed one after another, and the spectral range has been greatly extended.